Andrzej Chwojnowski

Spongy Polyethersulfone Membrane for Hepatocyte Cultivation: Studies on Human Hepatoma C3A Cells

There are different types of membranes used for hepatocyte cultivation. In our studies, spongy polyethersulfone (PES) membranes were examined as a support for hepatic cell cultivation in vitro. The extended surface of the membranes allows to introduce a high cell number especially in three-dimensional gel structure. Scanning electron microscopy analysis indicated that C3A cells used in our experiments grew well on PES membranes forming microvilli characteristic for normal hepatocytes. Analysis of cell viability proved that spongy PES membrane is well tolerated by J774 macrophages and did not stimulate nitric oxide synthesis. Bile canalicular structures were observed in fluorescence microscopy after F-actin staining with tetramethyl rhodamine iso-thiocyanate (TRITC)-phalloidin. The C3A cells showed high affinity to the PES membranes and adhered to almost 90% during the initial 24 h of incubation. Albumin production increased during static culture from the value of 805.2 +/- 284.4 (ng/24 h/initial 10(6) cells) during the first days, to 2017.6 +/- 505.9 (ng/24 h/initial 10(6) cells) after 10 days of culture. In conclusion, the spongy PES membranes can be used as scaffold for hepatocyte cultivation, especially for the creation of three-dimensional environments.

Electrostatic Droplet Generator with 3-Coaxial-Nozzle Head for Microencapsulation of Living Cells in Hydrogel Covered by Synthetic Polymer Membranes

The current methods allow for encapsulation of cells inside spherical microcapsules made of a matrix covered by a permselective membrane using an electrostatic droplet generator with 1-nozzle or 2-nozzle heads. However, some potentially useful materials for the outer membranes cannot be put into direct contact with hydrophilic core filled by cells during the manufacturing process. Therefore, we designed a novel 3-coaxial-nozzle head that allows for the third fluid to separate the core material from the membrane material. The equipment was applied for manufacturing spherical microcapsules comprised of cell-friendly alginate core surrounded by semipermeable polyethersulfone membrane. The obtained microcapsules had a diameter between 0.84 mm and 1.79 mm, and the diameter correlated negatively with the applied electric voltage. The thickness of the membrane varied from 171 µm to 450 µm. The SEM images of the interior of microcapsules revealed highly porous membrane structure typical for synthetic membranes obtained by a wet phase inversion method. Bakery yeast cells encapsulated inside the alginate-polyethersulfone microcapsules retained their proliferation ability proving the effectiveness and safety of this encapsulation technique.

Transmembrane Pressure as an Indicator of a Density of Endothelial Cells Cultured Inside Capillaries of a Membrane Bioreactor under Dynamic Conditions

The main objective of this study was to asses usefulness of usage of transmembrane pressure changes measurements for monitoring of cells culture in the closed construction of bioreactor with semipermeable capillary membranes. Two simultaneous HUVECs’ cultures in bioreactors with an assumed cells densities of 20,000 and 60,000 cells/cm2 were carried out. Obtained results indicated that the transmembrane pressure rise time of 30 mmHg after locking up the capillaries exit depend on the quantity of cells introduced for seeding to inner part of capillaries and may be a good marker of cells quantity cultured on the inside surface of capillary membranes. However microscopy analysis of the inner part of membranes after end of cultures showed that cells shapes and sizes differed from those for cells cultured without monitoring of transmembrane pressure changes. The further studies are necessary aimed at selection of the value of the transmembrane pressure rise which does not influence on the cells morphology, cytoskeletal organization and does not disturb physiological behavior of the cells.

Semi-permeable polysulfonic membranes to obtain dry tests

Abstract A method of covering a cellulose support with a semi-permeable membrane is discussed. This covering was carried out in a continuous manner with very good reproducibility. Due to the expected application and specific properties of the membrane obtained, a non-standard method of evaluation of the quality of the material appeared to be necessary. An objective method of studying the permeation time of the permeate through the membrane deposited on the support was determined. All the membranes obtained were studied utilizing this method. The weight composition of the membrane-forming mixture, the parameter which had an effect on the permeation time, was studied. The membraneforming polymer to pore-forming agent weight ratio and the pore-forming agent molecular weight were studied. The relationship between the times of depositing the membrane-forming mixtures and permeation times was also established. It was found that in the membranes obtained, no pore collapsing occurred.

Computer-aided analysis of micro-morphological structure of porous membranes

BackgroundThe paper presents an approach and computer-aided method of numerical evaluation of the quality of porous membranes used as scaffolds for cultivation of chondrocytes in regeneration of biological tissues.MaterialsThe scanning electron microscope (SEM) images of 300× and 1000× magnification presenting the sections of artificial polyvinylpyrrolidone membranes obtained in two alternative production processes are examined.Theory and methodsThere is presented a combined morphological and statistical method of the assessment of artificial membranes’ porosity, based on computer-aided segmentation and analysis of the size and shape of pores. Theoretical backgrounds of description pores as irregular objects in discrete 3-dimensional space are presented. The parameters characterizing the quality of pores: pores irregularity coefficient and pores density are defined. The quality of the examined specimens of materials is characterized by the size (mean 2-dimensional section areas) of pores. The main concept presented in the paper is the extraction of lacking information concerning the third dimension of pores from the 2-dimensional SEM images of their sections. Two approaches to evaluation of the parameters characterizing pores on the basis of computer-aided analysis of their cross-sections are proposed: (1) based on statistical extension of geometrical data and (2) based on analysis of brightness profiles. The corresponding methods are based on the assumption of isotropy of the examined porous materials. The results of automatic measurements of the areas of pores, lengths of their chords and recording the brightness profiles along fixed lines crossing the analyzed images are illustrated by examples.ConclusionsPractical usefulness of the proposed methods to evaluation of the quality of porous membranes consists in their ability to be used in case if alternative methods for some reasons cannot be used.

Polysulfone/polyurethane blend degradable hollow fiber membranes preparation and transport–separation properties evaluation

AbstractThe method of preparation of partly degradable polysulfone/polyurethane (PSf/PU) asymmetric hollow fiber membranes is presented. The membranes built of PSf and polyurethane polymer containing degradable ester groups were obtained by dry/wet-spinning phase-inversion technique then treated with sodium hydroxide solution using the flowing method. The membrane hydraulic permeability (UFC) for pure water and retention coefficients were evaluated for chosen markers before and after applied hydrolysis. An increase in ultrafiltration coefficient was observed while retention coefficient did not change significantly in post-treatment membranes. Hydrolysis caused the partial removal of PU from the membrane structure resulting in the opening of the pore passages and the permeability increase without increasing the pore size. The morphology of the PSf/PU hollow fiber membranes was analyzed using scanning electron microscopy before and after hydrolysis.